Jamal and colleagues recently published their results from a randomized, controlled trial on the effects of nitroglycerin ointment on bone density and strength [1]. This study follows several previous ones [2,3] which showed that the use of nitrates had a positive impact on bone mineral density (BMD) in men and women. 

The present cohort included 243 postmenopausal women (who were at least 1 year postmenopausal), mean age 62 years, with BMD T-scores between 0 and -2.0 at the lumbar spine. The daily dose was 15 mg (2% nitroglycerin ointment) applied at bedtime to their upper outer arm. The intake of calcium was 1200–1500 mg/day and that of vitamin D was 800 IU/day from food and provided supplements. The duration of treatment was 24 months, and the study ran from November 2005 until March 2010. The primary endpoint was BMD T-score measured at 12 and 24 months, at the lumbar spine, femoral neck and total hip. At 24 months, 118 of 126 women in the nitroglycerin group and 111 of 117 women in the placebo group completed all the assessments. 

At 24 months, spine and femoral neck BMD significantly increased by 6–7% in the nitroglycerin-treated group compared with placebo: lumbar spine BMD increased from 1.05 g/cm2 to 1.14 g/cm2, vs. 1.06 g/cm2 to 1.08 g/cm2, respectively. The corresponding figures for total hip were 0.92 to 0.97 g/cm2 vs. 0.93 to 0.92 g/cm2, and for femoral neck 0.88 to 0.93 g/cm2 vs. 0.87 to 0.86 g/cm2; p = 0.001). Cortical bone parameters assessed using peripheral quantitative computerized tomography also demonstrated an increase in volumetric trabecular BMD, cortical thickness and periosteal circumference at the radius and tibia compared with placebo. Urine N-telopeptide, the marker for bone resorption, decreased by 54%, while bone-specific alkaline phosphatase, the marker for bone formation, increased by 35% in women treated with nitroglycerin. There were two fractures in both the treated and the placebo groups, and one death by stroke in the treated group. The adverse events in the active group were dominated by headaches and were frequent at the beginning of the treatment, but rarer as a new event in the second year. The conclusions of the study were that daily administration of nitroglycerin ointment increased bone formation and decreased bone resorption, improved BMD, bone structure and indices of bone strengths ‘at least as much as existing treatments’ and also ‘may reduce the risk of vertebral fractures and non-vertebral fractures’.

Comment
In March 1910, JAMA published an article on the effects of nitroglycerin in workers of the explosives industry, calling them ‘glycerin men’ or ‘powder men’. It was revealed that nitroglycerin may enter the human body through the skin, respiratory system and alimentary tracts, leading to immediate side-effects such as headache, nausea and vomiting, as well as some long-term ones like shortness of breath. It was noticed that symptoms might disappear with time if the exposure was removed, but could recur after returning to work [4].

Apparently, the headaches are still the main problem and side-effect of nitric oxide (NO) donors used as medication. The trial by Jamal and colleagues was preceded by a 1-week run-in phase in which all the patients received active substance – ointment 2% with nitroglycerin. From the 400 women enrolled (eligible by study criteria), just 243 remained in the study, and from these 104 discontinued the study because of headaches, or headaches with nausea, and two because of allergic reactions, giving a drop-out rate of 25%. Headache was mentioned by all the previous relevant bone trials as being the main problem and reason for early withdrawal.

The results of the study by Jamal and colleagues [1] were in contrast to the NOVEL Study (The Nitroglycerin as an Option: Value in Early Bone Loss) published by Wimalawansa and colleagues [5] in 2009. NOVEL was a 3-year, open-label study on 186 subjects of whom half were in the treated group and half in the placebo group. The study concluded that BMD changes did not substantially differ between postmenopausal women who received once-daily dosing of 22.5 mg transdermal nitroglycerin and those on placebo. Note that the compliance adjusted dose was only ~16 mg/day, a sub-therapeutic dose. 

Rejnmark and colleagues conducted a larger study on nitrate usage and bone effects [6] – a population-based, pharmaco-epidemiological, case–control study, which included 124,665 persons who sustained a fracture in comparison with a group of 373,963 controls matched for age and sex. The statistical analysis showed, after adjustment for confounders, that use of nitrates (isosorbide mononitrate, isosorbide dinitrate, and nitroglycerin) during the previous 5 years was associated with a 11% reduced risk of any fracture (odds ratio (OR) 0.89; 95% confidence interval (CI) 0.86–0.92) and a 15% reduced risk of hip fracture (OR 0.85; 95% CI 0.79–0.92). Risk of any fracture was reduced in both men and women, but risk of hip fracture was only reduced in women. Hip fracture risk was reduced dose-dependently. Use of nitrates with a short duration of action was associated with lower risk estimates than use of slow-release preparations [6].

The effects of the NO donors on the bone might be influenced by several factors.
• The mechanism of action is partly mediated through the NO/cyclic guanosine monophosphate pathway and may also be mediated via IGF-I. Postmenopausal estrogen deficiency is associated with cytokine-induced increases in bone resorption. Low levels of nitric oxide are associated with similar cytokine-induced activity on bone, and high levels of NO block this cytokine-mediated increase in bone resorption. Furthermore, menopause is associated with low serum levels of NO, which are increased by estrogen replacement therapy. Thus, NO deficiency may mediate the bone loss associated with menopause, and this may be prevented by the administration of NO donors [1].
• Doses may vary: across many studies, intermediate doses of NO have been shown to promote skeletal health, whereas high doses, such as are used in angina pectoris, may promote bone loss. Low doses of NO have suppressed osteoclast bone resorption and promoted growth of osteoblasts in vitro. Lower concentrations of NO stimulate osteoblast and osteocyte activity and control osteoclast-mediated bone resorption. Higher doses lead to bone loss through osteoclast activation and osteoblast suppression [5].
• Tachyphylaxis has effects on receptors, which were suggested by the superiority of the intermittent administration of gel compared with daily use or multiple use [1].
• Bioavailability also may vary between different NO donors and nitrate preparations [7].
• The therapeutic window for treatment of osteoporosis is relatively narrow and there is a relative need for a higher dose in early menopause (because the bone turnover is relatively high during the early postmenopausal period, it is possible that the dose of nitroglycerin required to prevent bone loss in this group of women could be higher). This was suggested to be one of the reasons for the negative results in the study by Wimalawansa and colleagues [5] vs. the good results in the study by Jamal and colleagues [1,3].
• Vascular effects should also be taken in consideration, near the classical mechanisms as used in osteoporosis therapies.
• There are associated pathologies in the given age group.

The results of the study by Jamal and colleagues [1] are very interesting and highlight a potential new benefit for an old, relatively cheap medication which is used worldwide. The data in this study do not have the statistical power to assess the fracture prevention (only two fractures in both groups), but the effects on cortical bone, as demonstrated by radius and tibia measurements, were promising in regard to possible non-vertebral fracture prevention. These changes in cortical bone parameters are greater than those observed with current antiresorptive medication or teriparatide [3]. Clearly, NO donors might be a new direction for clinical research and indication in osteoporosis, and a possibility for individualization of the treatment in difficult cases with multiple pathologies.

Iuliana Ceausu
Senior Lecturer, Obstetrics and Gynecology, ‘Carol Davila’ University of Medicine and Pharmacy, and Senior Specialist in Obstetrics and Gynecology, Department of Obstetrics and Gynecology, ‘Dr. I. Cantacuzino’ Hospital, Bucharest, Romania

References
1. Jamal SA, Hamilton CJ, Eastell R, Cummings SR. Effect of nitroglycerin ointment on bone density and strength in postmenopausal women: a randomized trial. JAMA 2011;305:800-7.
http://www.ncbi.nlm.nih.gov/pubmed/21343579
2. Jamal SA, Browner WS, Bauer DC, Cummings SR. Intermittent use of nitrates increases bone mineral density: the study of osteoporotic fractures. J Bone Miner Res 1998;13:1755-9.
http://www.ncbi.nlm.nih.gov/pubmed/9797485
3. Jamal SA, Goltzman D, Hanley DA, et al. Nitrate use and changes in bone mineral density: the Canadian Multicentre Osteoporosis Study. Osteoporos Int 2009;20:737-44.
http://www.ncbi.nlm.nih.gov/pubmed/18800179
4. Laws CE, Du Pont W. Nitroglycerin head. JAMA 1910;54:73, reprinted in JAMA March 3, 2010;303:891
5. Wimalawansa SJ, Grimes JP, Wilson AC, Hoover DR. Transdermal nitroglycerin therapy may not prevent early postmenopausal bone loss. J Clin Endocrinol Metab 2009;94:3356-64.
http://www.ncbi.nlm.nih.gov/pubmed/19549739
6. Rejnmark L, Vestergaard P, Mosekilde L. Decreased fracture risk in users of organic nitrates: a nationwide case-control study. J Bone Miner Res 2006;21:1811-17.
http://www.ncbi.nlm.nih.gov/pubmed/17054422
7. Khosla S. Is nitroglycerin a novel and inexpensive treatment for osteoporosis? JAMA 2011;305:826-7.
http://www.ncbi.nlm.nih.gov/pubmed/21343584